The proposed research project is focused upon the following areas of prostatic development all of which have relevance to proliferative diseases of the prostate such as benign prostatic hyperplasia and prostatic adenocarcinoma: (1) Analysis of mesenchymal differentiation during prostatic development; (2) Analysis of prostatic epithelial stem cells; (3) Analysis of regional heterogeneity within the normal and abnormal prostate; (4) Analysis of androgen-dependent ductal branching morphogenesis; (5) Analysis of epithelial-stromal interactions in hormonal imprinting leading to prostatic pathogenesis; and (6) Analysis of the mechanism of mesenchyme-induced regulation of epithelial proliferation. Mesenchymal differentiation and the emergence of smooth muscle cells during normal and abnormal prostatic development will be studied by morphometry and immunocytochemistry. Proliferative activity of urogenital sinus mesenchyme and the localization of extracellular matrix components in the mesenchyme will be assessed by autoradiography and immunocytochemistry. Prostatic epithelial stem cells will be studied by grafting tissue recombinants composed of urogenital sinus mesenchyme (UGM) and small fragments of prostate ducts containing 50 to 500 epithelial cells and determining their proliferative potential. Regional heterogeneity within the prostate will be assessed morphologically by light and electron microscopy, 3H-thymidine autoradiography (ARG), 3H-DHT ARG, and by immunocytochemistry of secretory proteins. Particular attention will be focused upon whether differences in biological activity correlate with androgen receptor activity. A new in vitro model of androgen-induced branching morphogenesis will be pursued utilizing cultures of bulbourethral glands (BUGs) growing in serum-free medium. Cell lineage studies will follow the descendents of single cells microinjected with non-toxic vital tracer dyes. Branching morphogenesis in the developing BUG will be examined specifically to assess the role of extracellular matrix components by histochemistry, immunocytochemistry, and ARG. The role of mesenchymal-epithelial interactions will be examined during the expression of abnormal prostatic epithelial development and growth elicited by neonatal imprinting with exogenous estrogen. Finally, the mechanism of mesenchymal regulation of epithelial proliferation in the BUG will be assessed in transfilter assemblies in a serum-free in vitro system to determine whether diffusible mediators are involved. Ultimately, mesenchymal cell conditioned medium experiments are planned.